The allele-specific PCR approach has been modified by introducing a second mismatch at the 3'-penultimate link of the primer and used to identify the sickle cell anemia mutation (A-->T transversion in the sixth codon of the human beta-globin gene causing Glu-->Val substitution in the protein), thus obviating the problem of an interpretationally ambiguous 3'-terminal mismatch including T residue.
The model used in this study is the amplification of a 725 base-pair (bp) beta-globin gene sequence encompassing the sickle-cell anemia point mutation, followed by Cvn I digestion.
The detection of the single base pair mutations at codon 6 of the beta-globin gene is important for the prenatal diagnosis of sickle-cell anaemia and SC disease.
The clinical diversity of sickle cell anemia is strongly related to the degree of intracellular hemoglobin S (Hb S) polymerization, which in turn is dependent on the intracellular concentration of Hb S. We have recently defined a region of DNA approximately 500 bp 5' to the human beta-globin gene that acts as a silencer for the transcription of this gene and have shown that a polymorphism in this sequence is associated with a thalassemic phenotype of the beta-globin gene.
It is likely that determinants unrelated to haplotype, linked or unlinked to the beta-globin gene cluster, are the major effectors of differences in the levels of HbF in American patients with sickle cell anemia.
A single base-pair mutation (beta s) in codon 6 of the human beta-globin gene, causing a single amino-acid substitution, is the cause of sickle cell anaemia.
A distinctive DNA polymorphism haplotype in the beta globin gene cluster (++- +-), tightly coupled to a C----T substitution at position -158 5' to the cap site of the G gamma globin gene, is strongly associated with sickle cell disease in this region.
Treatment of sickle cell anemia with 5-azacytidine results in increased fetal hemoglobin production and is associated with nonrandom hypomethylation of DNA around the gamma-delta-beta-globin gene complex.
Each pG gamma F is linked with one of the major haplotypes of the beta-globin gene cluster observed in sickle cell disease (SCD) associated with different mean levels of hemoglobin F (Hb F) expression (P < .001).
Implementation of this approach for disorders resulting from mutations affecting the beta-globin gene (e.g., beta-thalassemia and sickle cell anemia), however, has been hampered by the inability to generate recombinant viruses able to efficiently and faithfully transmit the necessary sequences for appropriate gene expression.
The molecular defect in sickle cell disease resides in the beta globin gene, with consequent defects in erythrocytes only, suggesting that the vascular occlusion and vasomotor instability which characterize this disease are the result of interactions between abnormal sickle erythrocytes and cells of the blood vessel wall.
Gene therapy approaches for beta-thalassemia and sickle cell anemia focus on the transfer of a human beta-globin gene into the patient's hematopoietic stem cells (HSC).
Our studies on the Saudi population from different regions of the country using Hinc II and Hind III showed that the beta-globin gene haplotype ++-++ is associated with a mild sickle cell anaemia, while ----+ is associated with the severe form of the disease.
This new generation of globin retroviral vectors may prove useful for gene therapy of human beta-globin gene disorders such as sickle cell disease and beta-thalassemia.
The inheritance of sickle-cell anemia upon the background of the major beta-globin gene cluster haplotypes has been associated with differing risks for major organ failure, and more recently with response to hydroxyurea treatment.